The present invention relates to the molecular cloning of cDNA for both A and B chains of human (hu) p53-specific, HLA restricted murine (mu) T-cell receptor (TCR), transfer of the cDNA to hu T cells, and functional expression of the p53-specific TCR in hu cytotoxic T lymphocytes (CTLs). The functional expression of the mu TCR results in the recognition of endogenously processed hu p53 expressed in tumor cells. The invention thus also relates to an anti-cancer immunotherapy by the adoptive transfer of TCR gene modified autologous T cells.
The publications and other materials used herein to illuminate the background of the invention or provide additional details respecting the practice, are incorporated by reference.
p53 acts as a checkpoint for cell division in most eukaryotic cells, and is normally expressed at low levels with rapid decay kinetics. Mutations in p53 characterize a large proportion of human (hu) solid tumors. These mutations, which abrogate the function of p53 as a suppresser of cell division (Finlay et al., Cell, 57:1083-1093, 1989) are associated with a prolonged half life and much higher nuclear and cytoplasmic concentration of the p53 protein (Eliyahu et al., Proc Natl Acad Sci USA, 86:8763-8767, 1989). The mutations are generally single base missense mutations (Kovach et al., Proc Natl Acad Sci USA, 93:1093-1096, 1996 ID), and the remainder of the over-expressed p53 molecule is wild type (wt). Unfortunately, the immune response to most tumors which aberrantly express p53 is not adequate to prevent their un-restricted growth. Either tolerance or other immune-based selective mechanisms may be responsible for inadequate host defense against these tumors.
Because the p53 gene product is over-expressed in a large proportion of all solid tumors, it provides an ideal target for enhancement of the T lymphocyte anti-cancer immune response. What makes wt p53 epitopes such an attractive target for an adaptive immune response that the intracellular concentration of p53 is normally very low (Zambetti et al., FASEB J., 7:855-865, 1993), and cells expressing normal p53 at low levels will most likely escape an enhanced immune response to over-expressed p53.
Murine (mu) experimental models have clearly demonstrated the ability to target over-expressed p53 as a means of achieving tumor rejection (Roth et al., Proc Natl Acad Sci USA, 93:4781-4786, 1996) 1996; Mayordomo et al., J. Exp. Med., 183:1357-1365, 1996). Furthermore, they have established that tumor rejection can be achieved without autoreactivity to cells that express normal levels of p53 (Vierboom et al., J. Exp. Med., 186:695-704, 1997). It remains a challenge to translate these findings into an effective immunotherapy strategy for hu malignancy because of the relatively poor hu immune response to p53-overexpressing tumors (Maehara et al., Cancer, Phila. 85:302-308, 1999). By contrast, mice generate a vigorous response to both mutant and nonmutant p53 epitopes (Vierboom et al., supra; Nijman et al., 1997; Bertholet et al., Eur. J. Immunol., 27:798-801, 1997).
The T cell receptor (TCR) is the surface molecule on T cells that recognizes processed antigen, either self or allo forms. The A/B form of the TCR has been recognized as the main form that recognizes alloantigen, in contrast to the G/D form, and it initiates a cascade in which cells expressing the recognized antigen are cytolytically attacked or growth restricted by the actions of T cell-secreted lymphokines. It has been shown that adoptive immunotherapy using A/B TCR expressing cytotoxic T cells will augment the eradication of tumor in a SCID model (McCarty et al., Cancer Res., 58:2601-2605, 1998; Stenholm et al., Int. J. Cancer, 77:476-480, 1998). The capability of CTLs to cause tumor regression is thought to be far greater than a humoral response or a CD4+ mediated Th infiltration of the tumor. Harnessing of the powerful anti-tumor property of CTLs has been elusive over the past 10 years; however, using gene transfer strategies with chimeric single chain TCRs provides a new approach that might overcome some of the earlier difficulties (Chung et al., Proc Natl Acad Sci USA, 91:12654-12658, 1994).
Previous studies (Yu et al., J. Surg. Res. 69:337-343, 1997) and by others (Theobald et al., Proc Natl Acad Sci USA, 92:11993-11997, 1995) have focused on immunizing transgenic (Tg) mice that express the hu transplantation antigen HLA A*0201 with immunogenic epitopes derived from wt hu p53. A potent CTL response was generated against one of these epitopes (Yu et al., supra). Isolated clonal CTLs selectively lyse/kill p53-overexpressing hu tumor cells in vitro and in in vivo tumor models in severe combined immunodeficiency (SCID) mice (McCarty et al., supra).
Consequently, it is desired to overcome the weak hu immune response to p53 by adapting to hu T cells the favorable characteristics of the powerful mu immune response to p53. It is further desired to develop compositions and methods for treating individuals having tumors which overexpress p53.
The present invention provides for the molecular cloning of cDNA for both A and B chains of hu p53-specific, HLA restricted mu TCR, the transfer of the cDNA to hu T cells, and the functional expression of the p53-specific TCR in hu CTLs. The functional expression of the mu TCR results in the recognition of endogenously processed hu p53 expressed in tumor cells. The invention thus also provides an anti-cancer immunotherapy by the adoptive transfer of TCR gene modified autologous T cells.